The PI has uncovered an essential Caulobacter gene, cgtA, that encodes a small monomeric GTP-binding protein and is a member of a recently identified subfamily of GTPases (the Obg family) conserved from humans to archea to bacteria. This evolutionary conservation between distantly related species suggests that this family of GTP-binding proteins shares an important physiological role. Obg proteins play a crucial role in cell growth, and in some cases, cell differentiation. Recently, it has been determined that the human homolog, DRG, functions in a pathway that is directly involved in oncogenesis in humans. Despite these recent advances, little is known about the mechanism by which these GTP-binding proteins function. Because of the ease with which cell cycle events in Caulobacter can be analyzed combined with the molecular and genetic advantages available when using a prokaryotic model system, they are in a unique position to uncover the cellular role of the Caulobacter GTPase, CgtA. Their long term goal is to elucidate the role of the CgtA mediated GTPase signal transduction pathway in Caulobacter. Towards this end, the laboratory has completed a molecular characterization of cgtA, has demonstrated that this gene is essential for viability, is a bona fide GTP-binding protein and is autophosphorylated. They have purified the protein and have raised CgtA-specific antibodies. They are now in a position to determine the mode of CgtA function, as well as the interplay between CgtA and as yet unknown interacting proteins involved in the same signaling pathway. During this funding period, they propose to: (1) isolate and examine both the in vitro GTP- binding characteristics and in vivo phenotype of specific cgtA mutants, (2) identify proteins that functionally or physically interact with CgtA, (3) determine the in vivo guanine nucleotide bond state of CgtA, (4) investigate the role of autophosphorylation, and (5) determine the intercellular location of CgtA by immunoelectron microscopy using antibodies they have generated to CgtA. From these studies, they will be able to suggest a role for this subfamily of GTPases and will develop tools which may be helpful for the identification of regulators and downstream effector proteins in other organisms.